Visual system neurons are highly dependent on oxidative metabolism for their normal functioning. When energy supply is diminished, visual neurons are impacted first and more severely than other neurons. Defective energy metabolism often leads to severe visual deficits, even blindness. Energy metabolism can be studied indirectly by monitoring increased blood flow and glucose utilization to areas of increased visual activity. Our laboratory studies it directly by analyzing the molecular mechanism of its regulation in visual cortical neurons. In the current grant period, we discovered that neuronal activity and energy metabolism are tightly coupled even at the transcriptional level. The same transcription factor, nuclear respiratory factor 1 (NRF-1), co-regulate genes for energy generation (exemplified by cytochrome c oxidase [COX], without which oxidative metabolism cannot be completed and mitochondrial ATP cannot be generated) and genes for glutamatergic transmission (NMDA receptor subunits 1 and 2B [Grin1 and Grin 2b], AMPA receptor subunit 2 [Gria2], and neuronal nitric oxide synthase [Nos1], a downstream mediator of NMDA receptor action) in visual cortical neurons. To probe the mechanism further, the present proposal has 3 specific aims to test our 3 hypotheses: (1) NRF-1 does not act alone, but rather in conjunction with another transcription factor, nuclear respiratory factor 2 (NRF-2), to co-regulate energy metabolism and neurochemicals of excitatory transmission in visual cortical neurons. We propose 3 models of operation: a) NRF-2 does not regulate any glutamatergic neurochemicals; b) NRF-2 regulates the same neurochemicals as NRF-1, providing a redundant mechanism; and c) NRF-2 regulates glutamatergic neurochemicals complementary to those of NRF-1, effecting a complementary mechanism. A combination of models 2 and 3 is also possible. This aim will be tested with multiple molecular biological approaches. (2) Even though NRF-1 and NRF-2 both regulate all COX subunit genes, they do not interact at the protein level. This parallel mechanism enables independent regulation by the two transcription factors. This aim and hypothesis will be tested with co-immunoprecipitation and mammalian two-hybrid system. (3) A tripartite mechanism exists by which the same transcription factor, NRF-1, regulates genes for energy generation (COX), energy consumption (Na+K+ATPase), and glutamatergic neurochemicals that mediate excitatory synaptic communication. Documentation of NRF-1's role in regulating Na+K+ATPase will provide the missing link and complete the circle of coupling between energy metabolism and neuronal activity. This aim and hypothesis will be tested with multiple molecular biological approaches as in aim 1. Results from these studies will significantly advance our understanding of the molecular mechanism by which energy generated exquisitely matches energy consumed by neuronal activity in visual cortical neuron. They will also provide the basis for future gene therapy for visual deficits resulting from faulty energy metabolism.